The present invention relates to a novel crosslinked polymeric substance having an amphoteric capability and furthermore relates to a process for preparing the novel crosslinked polymeric substance from natural chitin or regenerated chitin.
Fiberformes occurring in nature are divided broadly into collagen in higher animals, chitin in arthropods and in lower plants and cellulose in higher plants, and the frames of the living things are made by the sedimentation of apatite, calcium carbonate and lignin onto the above-mentioned fiberformes, respectively. Among them, chitin is a mucopolysaccharide of poly-N-acetyl-D-glucosamine, and its amount in nature is comparable favorably with the amount of cellulose in nature. However, since chitin is a highly crystalline substance and the intermolecular bonding thereof through the acetylamino groups is extremely stable, it is very much difficult to find an appropriate solvent to dissolve, disperse or swell chitin favorably. Accordingly, the development pertaining to the utilization of the chitin resources is far behind of that of cellulose and other polysaccharides.
The inventor of the present invention has formerly offered a method for preparing a water-soluble chitin-derivative (Japanese Patent Application No. Shou 53-161391/78) and a shaped material which makes most of the specific property of the water-soluble chitin (Japanese Patent Application No. Shou 53-161389/78). Particularly, the Japanese Patent Application No. Shou 53-161388/78 discloses a method by which variously shaped material having amphoteric ion-exchangeability has become available and contributes largely to the effective utilization of the chitin resource.
However, since in the outer surface of the above-mentioned shaped material, a larger part of amino groups have been acylated, the material is not amphoteric. Furthermore, since the membrane of the outer surface of the shaped material has a molecular sieve effect, it cannot allow the high-polymeric substance such as protein to permeate through the membrane into the inner part, and accordingly, there are cases where its amphoteric ion-exchangeability or its adsorbing capability are not sufficiently exhibited.
The inventor of the present invention, after studying earnestly, has developed a novel polymeric substance insoluble in aqueous solutions of acid, alkali and neutral salt, respectively, having carboxylic group(s) which may become anion and amino group(s) which may become cation, and a process for preparing of the novel polymeric substance and has been lead to the present invention.
The crosslinked polymeric substance of the present invention is synthesized from natural chitin or regenerated chitin as the raw material. The thus obtained crosslinked polymeric substance has (a) a degree of crosslinking of 0.01 to 0.3, (b) a degree of substitution of carboxylic group of 0.1 to 0.9 and (c) a degree of deacetylation of 0.1 to 1.
The following is the detailed description of the process for preparing the crosslinked polymeric substance according to the present invention.
Chitin used as the raw material of the present invention is the product obtained by a publicly known method from natural substances containing chitin represented by the following formula (I): ##STR1##
After crushing chitin, it is immersed into an aqueous alkaline solution of a normality of 10 to 15. The alkali used in the present invention is sodium hydroxide, potassium hydroxide or lithium hydroxide. The amount of the aqueous alkali solution is 3 to 5 times by weight of the amount of chitin, and the temperature of the aqueous solution is lower than room temperature, preferably of 5.degree. to 20.degree. C.
Then, in order to make the crosslinking of chitin easier, the excess alkali solution is removed from the immersed chitin, or the immersed chitin is subjected to freezing. Particularly, the freeze-treatment of the immersed chitin opens preferably the chitin micells, and is able to keep the alkali in the intricated parts of the micells, and accordingly, the treatment acts advantageously in the reaction of crosslinking.
In the freezing step, the mixture of chitin and the alkali is frozen at a temperature of -3.degree. to -30.degree. C., preferably of -10.degree. to -20.degree. C. for a preferable time of 3 to 24 hours. The degree of swelling the micells of chitin is determined by taking up a small amount of the specimen and dissolving it into a small amount of an iced water to see the uniformity of the thus obtained solution, because in the case of fully swelled micells, the solution is uniform with dispersed chitin.
In addition, in the cases where the amount of alkali is out of the above-mentioned range, the micells are only partially opened giving a non-uniform and opaque solution. The reason is considered that in cases of smaller amount of alkali solution, it cannot permeate into the interior of the chitin particles, and on the other hand, in cases of larger amount of alkali solution, it presents on the surface of the particle of chitin in plenty and only the surface is frozen in the freezing treatment. As a result, the interior swelling of the particles of chitin is suppressed not to open fully. In cases where the concentration of alkali is too small, chitin does not swell when immersed into such an alkali solution.
The crosslinking is carried out by admixing a crosslinking agent in an amount of 0.1 to 3 times, preferably 0.5 to 2 times by molecular weight of the raw material, chitin, with the above-mentioned chitin from which the excess of the aqueous alkali solution has been removed or with the above-mentioned alkali-containing chitin after treatment of freezing, at a temperature of lower than room temperature, preferably lower than 0.degree. C. for 5 to 48 hours.
The crosslinking agent is represented by the general formula: ##STR2## wherein X represents a halogen atom; Y represents an oxygen or a sulfur atom; Z represents a hydroxyl or a thiol group, and n is an integer of 1, 2 or 3.
An example of the crosslinking state of the crosslinked chitin obtained by the above-mentioned reaction is represented by the following general formula (II): ##STR3## wherein l, m and n are an integer.
Other than the structure shown in the formula (II), the above-mentioned crosslinked chitin are crosslinked via the divalent group ##STR4## between the oxygen bonded to 6-position of carbon of N-acetyl-D-glucosamine and the oxygen bonded to 3-position of carbon of the glucosamine, between the oxygen atoms respectively bonded to 6-positions of carbon atoms of two N-acetyl-D-glucosamines, or between the oxygen atoms respectively bonded to 3-positions of carbon atoms of two N-acetyl-D-glucosamines.
As is clearly seen in the above-mentioned structural formula, the crosslinking is carried out, as in the carboxyalkylation which is described later, on OH group or on CH.sub.2 OH group of chitin. Accordingly, the degree of crosslinking is determined in relation to the degree of carboxyalkylation. The degree of crosslinking in the present invention is 0.01 to 0.3 per unit of N-acetyl-D-glucosamine. The term "degree of crosslinking" is the reacted amount of crosslinking agent obtained from the values of elementary analysis. As the crosslinking agent represented by the formula ##STR5## for instance, epichlorhydrin, 1-bromo-3,4-epoxybutane, 1-bromo-4,5-epoxypentane, 1-chloro-2,3-epithiopropane, 1-bromo-2,3-epithiopropane, 1-bromo-3,4-epithiobutane, 1-bromo-4,5-epithiopentane, 2,3-dibromopropanol, 2,4-dibromobutanol, 2,5-dibromopentanol, 2,3-dibromopropanethiol, 2,4-dibromobutanethiol, 2,5-dibromopentanethiol, etc. can be mentioned, however, epichlorhydrine, 2,3-dibromopropanol and 1-chloro-2,3-epithiopropane are preferable in use.
In the next place, the crosslinked carboxyalkylchitin is obtained by the following method:
The above-mentioned alkali-containing crosslinked chitin is dispersed into an organic solvent containing an etherifying agent and made to react at a temperature of 0.degree. to 30.degree. C. for one to 72 hours, preferably 5 to 12 hours. In this case, since there is an evolution of heat due to neutralization at first, it is preferable to keep the temperature at 0.degree. to 10.degree. C. and then at a predetermined temperature in the reaction. After the reaction is over, the organic solvent is removed and the residue is dispersed into water. The aqueous dispersion is neutralized, filtered, desalted and dried to obtain the crosslinked carboxyalkylchitin. In the present invention, as the organic solvent in carboxyalkylation, a solvent selected from methanol, ethanol, isopropyl alcohol, n-propanol, acetone and dimethylsulfoxide is used. The reason why such an organic solvent is used, is that in the process of the present invention, since the hydroxyl groups of chitin take a form of alcoholate by the action of alkali, there is an advantage of possibly carry out the carboxyalkylation of the alkali-containing chitin as it is.
The etherifying agent used in carboxyalkylation in the present invention is a compound represented by the general formula of X(CH.sub.2).sub.n COOH, wherein X represents a chlorine or a bromine atom, and n is an integer of 1, 2 or 3, and is exemplified by chloroacetic acid, bromoacetic acid, beta-chloropropionic acid, beta-bromopropionic acid, gamma-chlorobutyric acid and gamma-bromobutyric acid.
The amount of the etherifying agent in the reaction is preferably one to three times by molecular weight of the raw material, chitin.
According to the process of the present invention, a water-insoluble derivative of chitin such as crosslinked carboxymethylchitin, crosslinked carboxyethylchitin and crosslinked carboxylpropylchitin is obtained. The degree of substitution with carboxyalkyl groups by the carboxyalkylation (hereinafter referred to as the degree of carboxyalkylating substitution) in the present invention can be determined at will by choosing the conditions, and it is generally 0.1 to 0.9, preferably 0.3 to 0.9. In addition, the degree of carboxyalkylating substitution, herein mentioned, is obtained from the elementary analytical data on the product and the titer of the salt-type chitin by an aqueous hydrochloric acid at the pK value of 4.3.
Furthermore, the crosslinked polymeric substance of the present invention comprising the de-acetylated product of carboxyalkylchitin is possibly obtained by de-acetylation of the above-mentioned crosslinked carboxyalkylchitin with an aqueous high-concentrated alkali solution, preferably an aqueous 4 to 15N alkali solution at a temperature of 65.degree. to 150.degree. C., preferably of 65.degree. to 110.degree. C. for 0.1 to 48 hours.
The above-mentioned crosslinked carboxyalkylchitin used as the starting material of de-acetylation may be the solvent-including product of the above-mentioned carboxyalkylation in an organic solvent obtained by filtering and washing with ethanol and acetone, the dried product or the product containing humidity.
During the reaction of de-acetylation, carboxyalkyl groups are not removed even when heated in the aqueous alkali solution because they are bonded by ether-linkage.
The degree of de-acetylation of the above-mentioned product of de-acetylation is obtained by the values of the elementary analysis of the product and is 0.1 to 1, preferably 0.3 to 1.
The properties of the crosslinked polymeric substance of the present invention obtained by the above-mentioned process have been confirmed by the elementary analysis, infrared spectroscopy, colour reactions, chemical analysis, tests for solubilities, etc. as follows:
(a) degree of crosslinking: 0.01 to 0.3; PA1 (b) degree of carboxyalkylating substitution: 0.1 to 0.9; PA1 (c) degree of de-acetylation: 0.1 to 1; PA1 (d) nitrogen content: 5.3 to 8.3% by weight; PA1 (e) infrared absorption bands: 1680 to 1720 cm.sup.-1 and 1500 to 1530 cm.sup.-1 ; PA1 (f) colour reaction: showing purple colour in reaction with iodine in the presence of sulfuric acid; and PA1 (g) solubility: insoluble in aqueous acid solutions, aqueous alkali solutions and aqueous salt solutions.
That is, in the infrared absorption spectrum of the object product of the present invention, an absorption band of 1680 to 1720 cm.sup.-1 corresponding to carboxyl group and an absorption band of 1500 to 1530 cm.sup.-1 corresponding to NH.sub.2 group are recognized and a purple colour is exhibited when the object product is made to react with iodine in the presence of sulfuric acid as a proof of a de-acetylated product. And the fact that the object product is crosslinked substance is verified by its insolubility in respective aqueous solution of acid, alkali and salts.
Moreover, the molecular weight of the crosslinked polymer having amphoteric ion-exchangeability, which the present invention offers, is not yet accurately determinable because there are no suitable solvents which are able to dissolve the polymer as it is (that is, without decomposition), as the raw material of the polymer, chitin, (because of the chemical stability of chitin, there are no solvents which are able to dissolve chitin as it is) although there are reports informing that the molecular weight of chitin is from a few hundred thousands to a few millions.